1. Field of the Invention
This invention generally relates to chemical reaction tube covers, and more particularly to a cover for a two-dimensional array of reaction tubes preferably utilized in an instrument for performing polymerase chain reactions (PCR).
2. Description of the Related Art
Automated thermal cyclers for performing PCR simultaneously on a number of samples are disclosed in the patent applications mentioned above and in U.S. Pat. No. 5,038,852. Briefly, PCR is an enzymatic process by which a small amount of specific DNA sequences can be greatly amplified in a relatively short period of time. The method utilizes two oligonucleotide primers that hybridize to opposite strands and flank the region of interest in the target DNA. A repetitive series of thermal cycles involving template denaturation, primer annealing, and the extension of the annealed primers by DNA polymerase results in the exponential accumulation of a specific DNA fragment whose termini are defined by the 5' ends of the primers.
A reaction mixture made up of the target DNA to be amplified, oligonucleotide primers, buffers, nucleotide triphosphates, and preferably a thermostable enzyme such as Taq polymerase, are combined and placed in reaction tubes. The reaction mixture contained in the tubes is then subjected to a number of thermal transition and soak periods known as PCR protocols in a thermal cycler to generate the amplified target DNA.
An array of reaction tubes is typically made up of up to either 48 or 96 tubes arranged in a 6.times.8 array or an 8.times.12 array in a tray. The array of tubes is placed in a metal thermal cycler block so that the lower portion of each tube is in intimate thermal contact with the block. The temperature of the block is then varied in accordance with the predetermined temperature/time profile of the PCR protocol for a predetermined number of cycles.
The denaturation step of the PCR protocol involves heating and maintaining the reaction mixture to around 95.degree. C. to separate double stranded DNA into single strands. At this elevated temperature, evaporation becomes a problem. To prevent evaporation of the tube contents during the PCR process, either a layer of wax or oil is placed on top of the mixture in each tube or a cap is placed on each tube in conjunction with a heated cover.
The caps are preferred over the oil or wax layer because application of such a layer is time consuming, messy, and invites mixture contamination. These caps may be separate individual caps or may be attached integrally to the tube. Alternatively, a series of plastic caps are connected together in linear strips of 8 or 12. Each one of the caps includes a tubular lower portion and an upwardly domed upper portion. The caps are connected together by an integral tab so as to form the strip of caps.
A tray of reaction tubes is typically filled with appropriate sample fluids, and each individual cap in a single strip is inserted into a tube so that the domed portion is up and the tubular portion fits down inside the reaction tube to provide a seal. The caps may be removed by pulling up on one end of the individual cap strip, as the reaction tubes are held within the tray by a retainer. Installation of these conventional caps on the reaction tubes is a relatively tedious and time consuming process requiring specific insertion of the tubular portion of each cap in each individual tube.
The tray of capped reaction tubes is inserted into a thermal cycler block and a heated platen cover is lowered over the block, pressing the domed caps downward to uniformly seat all of the reaction tubes and establish good thermal contact between each tube and the thermal cycler block. The heated platen cover provides a closed environment over the upper portions of the tubes projecting above the thermal cycler block. This heated platen cover is maintained during the thermal cycling protocol at a temperature greater than any of the thermal cycling temperatures so as to preclude vapor condensation within the upper portion of the tube or beneath the cap, both of which protrude above the body of the thermal cycler block. Thus, evaporative losses are prevented by the caps and internal vapor condensation is prevented by the elevated temperature under the platen cover.
The heated platen cover also prevents refluxing which affects the temperature of the sample within the reaction tube. Refluxing is the cyclical evaporation and condensation within the enclosed space above the sample within the reaction tube. Refluxing will generally lower the sample temperature during the thermal cycling protocol.
After the thermal cycling protocol has been completed, the tray of capped reaction tubes is removed from the thermal cycler and may be allowed to return to room temperature. The strips of caps are then removed from the tubes carefully so as to preclude cross-contamination between the tubes, and the array is transferred to other instruments for PCR product detection or further processing.
The configuration of plastic caps consisting of a strip of individual domed caps is quite adequate for small scale PCR where high throughputs are not required. The design offers the advantage of isolating each individual reaction tube but can be tedious to position in place and to remove. Accordingly, there is a need for a full plate cover or blanket which would offer the user an easier and faster way of sealing an entire array of tubes and easier, more efficient access to the tubes at the end of the PCR process.